new material - gene/cell therapeutic delivery Flashcards
(66 cards)
1
Q
Describe DNA therapeutics
A
- gene therapies
- therapy that is based on nucleic acid, resides in nucleus (center of cell)
- needs access to the nucleus to deliver and process genetic material
2
Q
Describe RNA therapeutics
A
- mRNA and sRNA therapies
- transcribed from DNA, delivered from cytoplasm (does not need access to the nucleus because translated in the cytoplasm, DNA -> RNA -> protein)
- easier to get into the cytoplasm compared to nucleus because travel time/space is much shorter
3
Q
Explain the difference between DNA & RNA therapeutics
A
DNA - not only do you have to cross the cell membrane, but you have to get to the nucleus
RNA - only need to cross the cell membrane
4
Q
Describe cell-based therapies
A
- they have a nucleus and a cytoplasm, which can be used to deliver genes in the DNA. but they can also act as their own therapies
- incorporate as new cells which perform cellular and metabolic functions
5
Q
Describe protein therapeutics
A
- protein antibodies
- can act extracellularly or intracellularly to affect a biotransformation or recognize a target
- in protein therapy, the active component is a protein (e.g antibody, hormone, etc), which acts on the cell surface, where antibodies will bind an antigen which is expressed on the surface of the cell
6
Q
Describe nucleic acid therapeutics
A
- any kind of RNA (mRNA, siRNA, etc.)
- they skipped the DNA step and have already been transcribed into RNA -> needs to go into cytoplasm
- NOT a gene therapy because RNA is TEMPORARY
7
Q
Explain why RNA is NOT gene therapy
A
- because RNA is TEMPORARY
- RNA leaves your body as soon as it is done with what it is supposed to do
ex - siRNA therapy will silence a gene, and then it will leave
8
Q
Explain the purpose of mRNA in the COVID vaccine
A
- To become a protein that can recognize the protein on the virus
- After it completes the “job,” the mRNA will either degrade or spin out
9
Q
Describe FDA-approved ASOs (drug substance)
A
single stranded DNA complementary to RNA
10
Q
Describe FDA-approved apatamers (drug substance)
A
single stranded DNA or RNA
11
Q
Describe FDA-approved RNAi (drug substance)
A
dsRNA
12
Q
Describe FDA-approved mRNA therapeutics (drug substance)
A
dsRNA
13
Q
Is a covid vaccine that delivers mRNA a gene therapy?
A
NO
it is a RNA therapeutic because it delivers mRNA to the nucleus to express a protein
14
Q
Is CAR-T a gene therapy?
A
YES
15
Q
Describe the location of action of CRISPR
A
It is DNA, needs to act all the way to the nucleus, in terms of being delivered
16
Q
Describe FDA-approved mAbs (drug substance)
A
protein
17
Q
What does “in vivo gene therapy” refer to?
A
AAV delivered DNA
18
Q
Describe “in vivo gene therapy”
A
treatment where the therapeutic agent (gene) or modified cells are directly introduced into the body to work inside and help treat disease
ex. vector-based therapies (FDA approved)
19
Q
Describe “ex vivo gene therapy”
A
treatment where cells are taken from the body, modified or tread outside in a lab, and then put back into the body of the patient to fight disease
**genetic modification happens outside body (ex: CAR-T, CRISPR) **
20
Q
Give an example of ex vivo therapy
A
blood draw. you extract blood out of the pt, you get the cells from the blood, you engineer the cells in the lab, you modify the cells, reinfuse in patient
(CELLS NEED TO COME BACK INTO BODY TO BE CONSIDERED EX VIVO)
21
Q
Describe oncolytic viruses (drug substance)
A
DNA
22
Q
Describe peptides (drug substance)
A
proteins
23
Q
List types of cell-based therapies
A
- stem cells & stem cell-derived products (hemopoietic) stem cell transplantation
- immunotherapies
24
Q
Define “TIL”
A
“tumor infiltrating lymphocytes”
cells associated with a tumor
25
Explain how TILs are used as a therapy
- Lymphocytes from a patient’s tumor that are extracted, amplified, and reintroduced to fight the cancer
- T-cells with multiple TCR clones, thus able to recognize shared antigens
- Contain effector memory T cells and T cell which express chemokine receptors for tumor honing
26
Define "TCR"
"T-cell receptors"
engineered cells based on cell-specific intracellular receptors
27
Define "CAR-T or CAR-NK"
"chimeric antigen receptor T- or NK- cells"
gene edited cells
28
Explain how TCRs are used as therapy
- gene edited cells (based on a real sequence of a real genetic interaction in the body)
- T cells are genetically engineered to recognize and attack cancer cells based on intracellular proteins
- Mostly T-cells
29
Explain the main difference between CAR-T & TCR
Main difference between TCRs and CARs is that TCRs are based on an naturally occurring interaction whereas a CAR is completely synthetic
30
Explain how CAR-T / CAR-NK is used as therapy
completely synthetic genetic construct (e.g antibody fragment) that allows the cell to make a protein, binding to a cancer cells and targeting a tumor antigen
31
How is stem cell therapy a thriving business in the US?
because stem cells can become healthy eye cells
32
Describe FDA-approved stem cell therapy products
hematopoietic stem cell transplantation from cord blood
33
Name indications for FDA-approved cell-based therapy products (CAR-T, TCR, or CRISPR)
blood disorders (cancers or sickle cell disease) & synovial sarcoma
34
Name the indication for FDA-approved cell therapies (TILs)
melanoma
35
FDA-approved cell-based therapies are also ______
immune cell therapies
36
List the main components of CAR
- Mostly made up of NK and T-cells
- Chimeric antigen receptors: synthetic genetic construct targeting a specific protein (antigen) on target (cancer) cells and deliver an activation signal to the cells
- Activation signals result in killing of the target and are given costimulatory domains, such as 4-1BB, CD28, CD8
- Designed to target tumor-specific antigens (TSAs) or tumor-associated antigens (TAAs)
37
Which of these cell therapies is or can be an immunotherapy?
A. Tumor infiltrating lymphocytes
B. CAR-T
C. Cancer vaccines
D. All of the above
D. All of the above
**everything is currently FDA approved**
38
Explain the basic components of CAR-T therapy
T-cells are removed, genetically engineered, expanded and delivered back to the patient:
- T-cells translates the gene into the protein
- Gene is synthetic, not made from patient’s DNA
- Expressed protein (CAR) is designed to recognize an antigen on cancer cells
Drug product: immune cell + gene (CAR-T/NK cell)
Drug substance: gene (DNA) expressed by the cell to become a cell-surface protein
T-cell is engineered with a DNA vector encoding the CAR protein. CAR is a protein that spans intracellularly.
39
Define "autologous cell-based therapy"
cells from your own body (you cannot donate to someone else)
similar to how people need to match between the donor and the recipient for blood transfers, but it’s more strict
Harder to source, patients are typically immunocompromised
40
Explain why cell donation for autologous cell-based therapy is more strict than blood transfers
it is more strict because there are T-cells that rejects infusion of things that are not their own
41
Is graft vs host disease (GvHD) a potential issue with autologous cell-based therapy?
typically, no
higher doses, feasible
42
Explain how genes are most commonly engineered into cells
VIRALLY
genes are most commonly engineered into immune cells virally using lentiviruses or retroviruses (for all FDA cell therapies)
43
Define "graft vs host disease (GvHD)"
- When patients infused with exogenous cells or transplanted with donor tissue recognize these transplants as foreign and react unfavorably
- Arises as a result of autoreactive (self-attacking) immune cells
44
Define "allogenic cell-based therapy"
cells from a different donor (universal T cells)
45
Give an advantage of allogenic cell-based therapy
Easier to source “off-the-shelf”
46
Give a potential issue from allogenic cell-based therapy
Can induce GvHD/immune response
47
Explain lymphodepletion
Lymphodepletion is a medical process involving the limitation/destruction of lymphocytes and T cells, typically through chemotherapy, to prepare the body for immunotherapy
48
Describe the purpose of lymphodepletion
purpose: disable immune system to avoid rejection
49
What drug combination is typically used for lymphodepletion?
Typically done with a combination of cyclophosphamide and fludarabine
50
What is the role of fludarabine
- Inhibits DNA polymerase (synthesis)
- Immune cells accumulate, the active metabolite of fludarabine, making them sensitive to lymphodepletion
51
All FDA-approved CAR-T cell therapies are ___ & ____
All FDA-approved CAR-T cell therapies are **autologous** and are **ex vivo**
52
What agent is commonly used for cryopreservation of CAR-T cells?
most common cryopreserving agent = DMSO
53
What receptor does Kymriah target?
CD19
54
List the potential risks of cell-based therapies
- sterilization of final product is not possible
- manufacturing is a long process, issue if patient is really sick
- cell viability requirements cannot always be met
- donor cells cannot always expand optimally
- complex logistics (temp control)
55
What antibody is used to commonly treat CRS?
IL-6 antibody (tocilizumab)
56
Explain how the risk of a non-sterile cell-based therapy product is prevented
You have to make the whole process is sterile by using aseptic processing key because you can’t sterilize only at the end
57
Which drug is used to commonly mitigate cytokine release syndrome (it is based on one cytokine only)?
Tocilizumab (antibody against IL-6)
58
Explain the role of the extracellular region of a CAR
- Responsible for binding specifically to an antigen present on the target cell
- Determines specificity of CAR-T cell by directing it to cells that express corresponding region
- Binding of extracellular region initiates activation cascade that leads to T-cell activation
59
What are cytokines (roles)?
- Proteins that activate and expand T-cells
- Proteins which support activation, maturation, and effector functions of immune cells
60
What are two problems that could arise when developing immunotherapies that target a single tumor antigen?
- On target, off tumor effects
- Fratricide
61
Describe on target, off tumor effects that can arise when developing immunotherapies that target a single tumor antigen
- Cells target correct antigen, but expressed on tissues other than target cancer
- Many cancer antigens also expressed on healthy tissues (albeit in smaller quantities)
- Can result in severe toxicities
62
Define "Fratricide"
Cells target antigen express on themselves
63
Define cytokine release syndrome (CRS)
Systemic inflammatory response, also called “cytokine storm”, that results in the release of a high number of pro-inflammatory cytokines that attack the patient’s immune system (leads to death)
64
What is a common target antigen for CAR therapy for leukemia?
CD19-expressed on blood cancer cells
65
What are two reasons why cell-based therapies have been more successful (and approved by the FDA) for blood cancers, but not solid tumors?
1. Immune cells cannot penetrate and kill large masses of tumors well
2. Solid tumors are highly immunosuppressive
66
What is one way to clinically mitigate or manage cytokine release syndrome?
Use of IL-6 antagonists (Etanercept and Toclizumab) that block IL-6 activation
